The insulin producing tissue of the pancreas, the islets of Langerhans, constitutes between about one and two percent of the mass of the pancreas. The isolation of the islets is desirable for laboratory purposes and for transplantation purposes. Transplantation of islets is looked to as a possible treatment for diabetes. Transplanting islets rather than segments of or an intact pancreas offers several advantages, including the ease of transplantation, the possibility of in vitro treatment to prevent rejection without immunosuppression, the elimination of the pancreatic exocrine function (the secretion of digestive substances from the host tissue), the possibility of cryopreservation of the tissue for subsequent use, and the possibility of xenografts.
The development and present state of the art in islet separation and transplantation is thoroughly reviewed in Volume 8, No. 2 of the World Journal of Surgery, April 1984, incorporated herein by reference. Previous work on separating islets of the rat pancreas is reported in The Journal of the American Diabetes Association, Vol. 16, No. 1, pp. 35-39, "Method For the Isolation of Intact islets of Langerhans From the Rat Pancreas," Lacy, et al., also incorporated by reference herein. In the original method of islet separation, still in use today, chopped pancreatic fragments are mixed with collagenase and incubated at 37.degree. C. until the maximum amount of islets are freed. The collagenase breaks down of digests the pancreas tissue, freeing the islets. The collagenase also acts on the islets, so that the islets released early in the process are broken down into single cells. If the process is stopped to protect the islets released early, many islets remain trapped in pancreatic fragments. Thus, only a fraction of the available intact islets are released by this method. This process is particularly ineffective for the isolation of islets from the pancreases of larger animals such as humans or dogs or pigs.
Laboratory islet isolation was greatly improved by the discovery that mechanical distension of rodent pancreas increased islet yield by causing mechanical separation of islets from the pancreas tissue. However, the same effect has not been noted in the pancreases of larger animals such as humans or dogs or pigs. After distension, the pancreas is chopped for collagenase digestion. This is presently accomplished by hand with scissors, which is unsatisfactory for mass processing, but mechanically powered chopping devices have been found to be too destructive to the islets.
Another variation of the collagenase isolation of islets is to wash the chopped pancreas fragments with an enzyme solution through a screen. This allows free islets to pass through the screen into a medium where they are protected from further enzyme activity.
Yet another technique employs Velcro.TM. fabric to hold the fibrous pancreas tissue while digesting it with the collagenase to release the islets.
All the present techniques for islet isolation, which are reviewed in Scharp, "Isolation and Transplantation of Islet Tissue," 8 World Journal of Surgery 143-151, 1984, incorporated herein by reference, are unsatisfactory for islet isolation, especially from the pancreases of larger animals. These techniques fall short in terms of efficiency of recovery and purity of the product. The procedures are simply inadequate for mass production techniques and the product is unsuitable for transplantation or other uses requiring purity.
The present invention is a new process for isolating the insulin producing tissue from a pancreas. The process involves ductal distension of the pancreas with a solution containing collagenase and incubating the pancreas at about body temperature, 37.degree. C. until a satisfactory separation of islets is observed. The pancreas is then quartered and macerated. The macerated pancreas is forced through successively smaller screens to filter out the pancreas tissue from the freed islets. The islet material can then be purified with an elutriator or other filtering means. The resulting product contains islet fragments and pancreas particles, as well as whole, viable islets, however, the islet content is sufficient to meet the requirements for transplantation purposes.
This process isolates viable, purified islets from human pancreases and from the pancreases of other animals. The process makes xenografts, the transplantation of islets of Langerhans from other animals to humans, possible because it removes most of the other pancreatic tissue and thereby reduces the chances of rejection of the transplant. Xenografts are particularly important because of the limited availability of healthy human pancreases for experimentation and transplantation.
The inventors have discovered that pig pancreases are very similar to human pancreases. This process can be utilized to isolate viable islets of Langerhans from pig pancreases for experiment and transplantation into humans.